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Identification of four insertion sites for foreign genes in a pseudorabies virus vector

文献类型： 外文期刊

作者： Zhang, Chuanjian 1 ; Guo, Shiqi 1 ; Guo, Rongli 4 ; Chen, Saisai 1 ; Zheng, Yating 1 ; Xu, Mengwei 1 ; Wang, Zhisheng; 1 ;

作者机构： 1.Jiangsu Acad Agr Sci, Natl Res Ctr Engn & Technol Vet Biol, Inst Vet Immunol & Engn, Minist Sci & Technol,Jiangsu Key Lab Food Qual &, Nanjing 210014, Jiangsu, Peoples R China

2.Jiangsu Coinnovat Ctr Prevent & Control Important, Yangzhou 225009, Jiangsu, Peoples R China

3.Nanjing Agr Univ, Coll Vet Med, Nanjing 210095, Jiangsu, Peoples R China

4.Jiangsu Acad Agr Sci, Inst Vet Med, Nanjing 210014, Jiangsu, Peoples R China

关键词： Pseudorabies virus; Bacterial artificial chromosome; Noncoding region; Insertion site; Spike gene

期刊名称：BMC VETERINARY RESEARCH （ 影响因子：2.741； 五年影响因子：2.955 ）

ISSN：

年卷期： 2021 年 17 卷 1 期

页码：

收录情况： SCI

摘要： Background: Pseudorabies virus (PRV) is a preferred vector for recombinant vaccine construction. Previously, we generated a TK&gE-deleted PRV (PRV Delta TK&gE-AH02) based on a virulent PRV AH02LA strain. It was shown to be safe for 1-day-old piglets with maternal PRV antibodies and 4 similar to 5 week-old PRV antibody negative piglets and provide rapid and 100 % protection in weaned pigs against lethal challenge with the PRV variant strain. It suggests that PRVTK&gE-AH02 may be a promising live vaccine vector for construction of recombinant vaccine in pigs. However, insertion site, as a main factor, may affect foreign gene expression. Results: In this study, we constructed four recombinant PRV-S bacterial artificial chromosomes (BACs) carrying the same spike (S) expression cassette of a variant porcine epidemic diarrhea virus strain in different noncoding regions (UL11-10, UL35-36, UL46-27 or US2-1) from AH02LA BAC with TK, gE and gI deletion. The successful expression of S gene (UL11-10, UL35-36 and UL46-27) in recombinant viruses was confirmed by virus rescue, PCR, real-time PCR and indirect immunofluorescence. We observed higher S gene mRNA expression level in swine testicular cells infected with PRV-S(UL11-10)Delta TK/gE and PRV-S(UL35-36)Delta TK/gE compared to that of PRV-S(UL46-27)Delta TK/gE at 6 h post infection (P < 0.05). Moreover, at 12 h post infection, cells infected with PRV-S(UL11-10)Delta TK/gE exhibited higher S gene mRNA expression than those infected with PRV-S(UL35-36)Delta TK/gE (P = 0.097) and PRV-S(UL46-27)Delta TK/gE (P < 0.05). Recovered vectored mutant PRV-S (UL11-10, UL35-36 and UL46-27) exhibited similar growth kinetics to the parental virus (PRV Delta TK&gE-AH02). Conclusions: This study focuses on identification of suitable sites for insertion of foreign genes in PRV genome, which laids a foundation for future development of recombinant PRV vaccines.

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